Acute myeloid leukemia (AML) is a devastating malignant disease of the hematopoietic system. Standard treatment for AML generally consists of cytarabine/anthracycline-based chemotherapy, however, only a minority of patients is cured with this approach. Thus, novel therapies targeting key leukemogenic pathways are needed. MicroRNAs (miRs) are short non-coding RNAs that regulate the expression of proteins involved in pivotal homeostatic mechanisms of normal and leukemia cells. In AML, deregulated miR expression is involved in disruption of hematopoietic mechanisms of cell differentiation, proliferation and survival, contributes to the molecular heterogeneity of the disease and impacts on treatment response and outcome. Among other miRs, we reported that higher expression of miR-155 predicts poor outcome, treatment resistance and is often associated with FLT3 internal tandem duplication (FLT3-ITD), a genetic marker predicting poor outcome in AML. However, miR-155 up regulation and prognostic impact is independent from FLT3-ITD and deregulation of this miR expression may represent a complementary hit for development of overt AML phenotype. We propose here to dissect the mechanisms through which miR-155 contributes to myeloid leukemogenesis and to explore how the therapeutic targeting of this miR may result in a clinical benefit in AML. We will accomplish this overarching goal through the following specific aims (SAs): SA1: To investigate the mechanisms through which miR-155 over-expression in the myeloid compartment contributes to leukemogenesis. We will develop conditional miR-155 transgenic murine models, where miR-155 will be expressed in different hematopoietic subpopulations, including stem cells and more committed myeloid progenitors. These mice will be characterized morphologically and functionally; if an overt leukemia phenotype is not observed, they will be crossed with the FLT3-ITD preleukemic mice; SA2: To determine the role and potential therapeutic benefit of targeting miR-155 in AML Leukemia Stem Cells (LSCs). We will dissect the mechanisms through which miR-155 potentiates LSC self-renewal and induces treatment resistance in AML; SA3: To investigate in vivo the pharmacokinetic (PK), pharmacodynamic (PD) and antileukemic activity of a synthetic LNA antimiR-155 using murine xenograft models of AML. We will conduct preclinical investigation of a novel, synthetic LNA antimiR-155 to evaluate the toxicity and PK and PD profiles and identify the active intracellular concentrations of the antimiR that induce efficient down-regulation of endogenous miR-155 and results in disease eradication in miR-155-dependent AML murine models; SA4: To target miR-155 in AML by conducting a phase I clinical trial of the neddylation inhibitor MLN4924 in combination with the hypomethylating agent (HMA) decitabine in relapsed/refractory AML patients. The rationale for this clinical trial is based on our data showing that MLN-4924 is a potent inhibitor of miR-155 expression and an ideal candidate to overcome miR-155-dependent mechanisms of treatment resistance.